PROJECT SUMMARY Bladder cancer represents a common malignancy worldwide. The main cause of death in bladder cancer patients is metastasis, with lymphatic metastasis as the primary means. Metastatic bladder cancer is currently difficult to remove completely, and not sensitive to radiotherapy and chemotherapy. Better understanding and exploitation of new diagnostic and therapeutic strategy for lymphatic metastasis of bladder cancer is urgently needed. The study of gene fusions has founded the theoretical backgrounds for many cancer diagnosis and therapeutics. However, even with whole genome sequencing, novel recurrent gene fusions are rarely identified in bladder cancer. Recently, our work on RNA trans-splicing and intergenic cis-splicing have helped open a new paradigm for intergenic splicing processes that generate chimeric RNAs with pro-tumorigenic activities, and may explain the molecular basis of the presence of driver onco-fusion products in the absence of chromosomal rearrangement at the DNA level. Indeed, in our preliminary studies, we have identified several chimeric RNAs unique to bladder cancer, but produced by intergenic splicing. Within a month since the issue of the RFA, together with our Chinese collaborators, we have successfully identified a few chimeric RNAs that are differentially expressed in lymph node metastatic group using only partial data. This success plus our past fruitful collaboration encouraged us to propose the following aims: 1. To identify of chimeric RNAs associated with lymphatic metastasis of bladder cancer by combining the RNA-Sequencing data from TCGA and China. By integrating with CPTAC and whole genome sequencing data associated with TCGA, we can determine whether the chimeric RNAs may code for chimeric protein, and whether they are products of gene fusion or intergenic splicing. Using the clinical information from TCGA and Chinese patients, we will reveal the relationship between chimeric RNAs and lymphatic metastasis, disease progression, and prognosis; 2. To investigate the role of the chimeric RNAs in lymphatic metastasis of bladder cancer cells, and to elucidate the molecular mechanism of the chimeric RNAs in regulating lymphatic metastasis of bladder cancer; We will focus on two groups of chimeric RNAs, fusion protein-coding and long non-coding chimeric RNA (lnccRNA). 3. To explore the significance of chimeric RNA as an early diagnostic marker, prognostic factor, and therapeutic target for lymphatic metastasis of bladder cancer. We will investigate the potential of the candidate chimeric RNAs as biomarkers, and nanoparticle delivery of siRNAs to target key chimeric RNAs; and 4) To develop an interactive web-based database to disseminate the results. The proposed study fits the NCI Funding Priorities, Genomics/Epigenomics/Transcriptomics/Proteomics, in that we will combine the large multi-omics data from both countries, analyze the landscape of chimeric RNAs, and investigate their roles as protein-coding, or long non-coding in lymph node metastasis of bladder cancer. !